a) Field of the Invention
The present invention relates to a process for the preparation of an N-protected .alpha.-L-aspartyl-L-phenylalanine methyl ester, which is an important intermediate compound for preparing .alpha.-L-aspartyl-L-phenylalanine methyl ester, which has sweetness of good quality and a sweetness degree of about 200 times the sweetness of sucrose. Therefore, its use as a diet sweetener is continuing to expand.
In the process of this invention, the N-protected .alpha.-L-aspartyl-L-phenylalanine methyl ester is prepared by the reaction of an N-protected L-aspartic anhydride with an acid addition salt of L-phenylalanine methyl ester in an organic solvent and in the presence either of a salt of an organic carboxylic acid or, when the organic solvent comprises an organic carboxylic acid, in the presence of an inorganic alkali metal or alkaline earth metal organic base, an ammonium alkali metal or alkaline earth metal salt of an organic carboxylic acid or ammonium carbonate.
The use of an acid addition salt rather than the free base form is critical for the prevention of self-cyclization reaction of the free amine in L-phenylalanine methyl ester and the ring cleaving reaction of the N-protected L-aspartic anhydride by water.
The process of the present invention lacks the disadvantages of the prior art, i.e., it avoids the production of byproducts by cyclization such as diketopiperazine compounds and it suppresses the production of .beta.-isomers, thereby improving the yield and purity of the end product. Moreover, when the reaction solvent comprises an organic carboxylic acid, the N-protected .alpha.-L-phenylalanine methyl ester can be selectively crystallized from the reaction mixture.
b) Description of the Prior Art
.alpha.-L-Aspartyl-L-phenylalanine methyl ester is a dipeptide compound composed of L-aspartic acid and L-phenylalanine methyl ester. Various methods have already been disclosed for the preparation of this compound and these methods are primarily chemical processes.
An N-protected L-aspartic anhydride is generally employed for the starting material. For example, in one known process, N-protected L-aspartic anhydride is subjected to a condensation reaction with L-phenylalanine methyl ester in an organic solvent and subsequently cleaving the protective group with a usual method to obtain .alpha.-L-aspartyl-L-phenylalanine methyl ester (U.S. Pat. No. 3,786,039).
In other processes, L-phenylalanine methyl ester is reacted with N-protected L-aspartic anhydride in an inert reaction medium. In accordance with the process of G.B. Application 708463, an ethyl acetate solution of N-benzyloxycarbonyl-L-aspartic anhydride and an ethyl acetate solution of L-phenylalanine methyl ester hydrochloride are mixed. A 1N aqueous sodium carbonate solution is added to the mixture to initiate the reaction by neutralizing the L-phenylalanine methyl ester hydrochloride salt.
In U.S. Pat. No. 4,824,994, L-phenylalanine methyl ester is reacted in a non-reactive solvent, e.g., toluene or toluene plus acetic acid, with N-carbobenzoxy-L-aspartic anhydride.
In a known process which does not use L-phenylalanine methyl ester, N-formyl-L-aspartic anhydride is subjected to a condensation reaction with L-phenylalanine in acetic acid and then the formyl group is removed in the presence of an aqueous hydrogen halide solution. The reaction product is subsequently esterified by treating with water, alcohol and the aqueous hydrogen halide solution, and .alpha.-L-aspartyl-L-phenylalanine methyl ester is isolated in the form of its hydrogen halide salt (U.S. Pat. No. 3,933,781).
A process for reacting N-benzyloxycarbonyl-L-aspartic anhydride with L-phenylalanine methyl ester in an inert organic solvent by reaction of the hydrochloride salt with a base is also known (Dahlmans et al., U.S. Pat. No. 3,808,190). For example, N-benzyloxycarbonyl-L-aspartic anhydride and L-phenyl-alanine methyl ester hydrochloride are dissolved in ethyl acetate and then an aqueous sodium carbonate or potassium carbonate solution is added to the resultant solution, which converts the hydrochloride acid addition salt of the L-phenyl-alanine methyl ester in situ into its free base form as evidenced by the disclosure in Dahlmans et al. at col. 2, lines 17-19, which states that the amount of base added is at least equivalent to the amount of the amino acid ester addition salt.
By adding a base at least equivalent to the amount of amino acid ester, the amino acid methyl ester acid addition salt is converted to its free base form, which is unstable and self-condensates to form a diketopiperidine. In fact, Dahlmans et al. disclose at col. 3, lines 13-15:
"the use of an aqueous solution of the base in the reaction of the present invention results in some racemization of the aspartyl-amino acid esters produced."
In contrast, the present invention is directed to the reaction of N-protected L-aspartic anhydride with a mineral acid addition salt of L-phenylalanine methyl ester and does not occur via the unstable free base form of L-phenylalanine methyl ester.
The reaction system of Dahlmans et al. contains an inert solvent, base and water. Although the inert solvent does not react with the reactants (see col. 4, lines 18-25), the base does and the adverse effect of using the free base form of the amino acid methyl ester inevitably occurs. In contrast, the reaction system of the present invention employs the stable form (acid addition salt) of the amino acid methyl ester. When the reaction of this invention is conducted in the presence of a base, the reaction solvent also includes an organic carboxylic acid, which is not merely an inert solvent because it prevents the formation of the unstable free base form of the amino acid methyl ester. Furthermore, no water is employed in applicants' reaction system, in contradistinction to Dahlmans et al. (col. 2, lines 22-24).
Takahashi et al. (U.S. Pat. No. 4,824,994) is directed to the production of N-carbobenzoxy alpha-L-aspartyl-L-phenylalanine methyl ester by reacting N-carbobenzoxy L-aspartic anhydride with L-phenylalanine methyl ester in its unstable free base form. The distinguishing feature of that process is the particle diameter of N-carbobenzoxy L-aspartic anhydride which has a particle diameter of not more than 130 .mu.m. Takahashi et al. discovered that the smaller the particle diameter, the higher the ratio of the .alpha. to .beta. isomers of N-carbobenzoxy alpha-L-aspartyl-L-phenyl-alanine methyl ester which is produced.
Takahashi et al. disclose at col. 2, lines 49-52, that the solvent is used merely for suspending the reactants. Although acetic acid is disclosed as an illustration of such an inert solvent, the acetic acid in Takahashi et al. is used merely as a suspending agent. Thus, as in Dahlmans et al., the amino acid ester is used in Takahashi et al. in a free base form, which is unstable under conventional conditions and which results in less than optimum yield and purity of the desired product.
Tertiary amines are also used in U.S. Pat. No. 3,808,190 as a base for promoting the reaction, in which case the reaction is carried out in the organic solvent by neutralizing the hydrochloride salt of L-phenylalanine methyl ester with an aqueous alkali solution. The inevitable disadvantage is that the ester group of L-phenylalanine methyl ester is hydrolyzed by the aqueous alkali solution, diketopiperazine compounds are formed as by-products by cyclization under alkaline conditions or basic conditions due to tertiary amines, in particular, and water causes ring-cleavage reaction of N-protected L-aspartic anhydride.
On the other hand, a process for conducting the reaction of N-formyl-L-aspartic anhydride with L-phenylalanine methyl ester in a solvent in the presence of acetic acid or formic acid is also known (EP 227301).
In U.S. Pat. No. 4,680,403, N-benzyloxycarbonyl-L-aspartic acid is converted to its anhydride in toluene with acetic anhydride and then reacted with L-phenylalanine methyl ester by the addition of a toluene solution thereof.
The L-phenylalanine methyl ester used as a raw material in these processes is usually obtained by reacting L-phenylalanine with methanol and a mineral acid such as hydrogen chloride, followed by neutralizing the resultant mineral acid salt of L-phenylalanine methyl ester.
However, the free amine form of L-phenylalanine methyl ester is thermally unstable and readily dimerized into diketopiperazine derivatives under conditions around neutrality, in particular, where neutralization and extraction operations are carried out. Additional limitations are placed on the yields of neutralization and extraction, and hence there is a disadvantage that the overall yield decreases on the basis of expensive L-phenylalanine.